Alloreactive T cells, central mediators of graft rejection, can recognize donor MHC: peptide complexes expressed on donor cells through the direct pathway of allorecognition and can recognize processed donor-derived peptide determinants expressed on recipient APCs in the context of recipient MHC molecules (the indirect pathway. While it is clear that CD4 + T cells responding through the indirect pathway participate in graft rejection, the presence and contribution of indirectly primed CD8 T cells to allograft rejection has not been addressed. We hypothesize that placement of an allogeneic transplant indirectly primes CD8 T cells, that these T cells comprise a significant portion of the indirect alloimmune response and that they migrate to the target organ. We further hypothesize that indirectly primed CD8 T cells contribute to the pathologic destruction of the transplanted organ through local effects at the graft site and by influencing the development of the remainder of the alloimmune T cell repertoire. We will test this hypothesis through the following specific aims. Aim 1. To compare the induction of the indirect CD8+ T call repertoire with the other components of the alloreactive T cell response to allografts. In these studies we will use complementary approaches to determine the contribution of indirectly primed CD8 T cells to the peripheral and intragraft alloimmune repertoire following skin or heart transplantation. Aim 2. To define the in vivo effector functions of indirectly primed CD8 T cells. In these studies we will use skin graft and heart transplant models to determine the in vivo effector functions and mechanisms of indirectly primed CD8 T cells and to fully assess their ability to mediate and/or contribute to graft pathology. Aim 3. To determine the in vivo requirements for indirect priming of CD8 T cells to transplant antigens. This aim will address the cellular and costimulatory requirements for priming CD8 T cells through the indirect pathway in vivo and will assess how eostimulatory blockade based interventions affect the function of these CD8 cells. The proposed studies will address an issue in transplantation immunology that has been ignored--whether and how indirectly primed CD8 T cells contribute to organ rejection. The design of effective therapies aimed at preventing rejection and improving human allograft survival depend on a complete understanding of the recipient alloimmune response. The findings derived from the work will define the role for indirectly primed CD8 T cells in allograft rejection and have the potential to guide future therapies aimed a prolonging graft survival.